Dr. Koval, is a senior fellow who will be promoted to a faculty position at the University of Rochester (UR). She has prior laboratory experience and a strong commitment to a career in academic Infectious Diseases. She seeks the funding to pursue 5 years of focused research on the pathogenesis of HIV-1 drug resistance. She wishes to build a strong foundation in research in order to make the transition to becoming an independent biomedical investigator. She will accomplish this through a co-ordinated training program incorporating course work, molecular biology and biochemical laboratory studies, and attendance at local and national conferences. She will be mentored by Dr. Lisa Demeter, and internationally recognized expert in HIV drug resistance, and Dr. Robert Bambara, a highly respected biochemist who has made fundamental contributions to understanding the mechanisms of HIV-1 reverse transcriptase (RT) function. Dr. Koval's research project will also have the support of the UR GCRC. The focus of Dr. Koval's studies will be to determine the pathogenic consequences of selecting for HIV-1 variants resistant to efavirenz (EFV), a potent non-nucleoside reverse transcriptase inhibitor (NNRTI). Resistance to EFV is unique among currently available NNRTIs, in that a primary RT mutation at codon 103 (K103N) develops first, followed by one of 3 secondary RT mutations (L100I, V108I, or P225H). The selective pressures favoring specific secondary resistance mutations is poorly understood. Dr. Demeter's laboratory has demonstrated that HIV variants resistant to other NNRTIs affect RNase H cleavage by RT. Variants with more extensive reductions in RNase H cleavage demonstrate reduced replication fitness in cell culture, and are less likely to develop in clinical HIV-1 isolates. We propose to further study whether improvements in RNase H cleavage and HIV-1 replication fitness can explain the selection for EFV-resistant (EFV-R) secondary mutations. We propose to pursue these important questions by characterizing: 1. The effects of secondary EFV-R mutations on HIV replication fitness of wild-type and K103N viruses; 2. The effects of secondary EFV-R mutations on wild-type and K103N RT function; and 3. The evolution of HIV-1 replication fitness in clinical isolates with increasing duration of EFV failure. In order to accomplish these specific aims, we will utilize a broad array of experimental approaches, ranging from biochemical studies of reverse transcriptase function, to analyses of HIV-1 replication fitness in patient samples.